S208: RELEVANCE OF DORMANT CANCER STEM CELLS IN DISEASE PROPAGATION AND THERAPY RESISTANCEIN JAK2V617F-DRIVEN MPN.
Authors: Ian Ghezzi, Esther Rodriguez Correa, Foteini Fotopoulou, Melanie Ball, Jeyan Jayarajan, Michael Milsom
Journal: HemaSphere (2023)

Background: Polycythemia Vera (PV) is initiated and maintained by mutant HSCs clones that acquired the JAK2V617F mutation and drive erythrocytosis in patients. So far, only Pegylated Interferon alpha (Peg-IFN-α) demonstrates a durable molecular response in the clinic; however, it is often associated with side effects leading to therapy discontinuation. Previous works suggest that Peg-IFN-α may deplete mutant HSCs by inducing their entry into cell cycle, DNA damage and subsequent exhaustion. This indicates that quiescence of mutant HSCs may be a vulnerable biological feature targeted by Peg-IFN-α. However, it has not been demonstrated that dormancy exists in the context of hematological malignancies, nor whether such cells are relevant to disease propagation and therapy resistance. Determining whether dormancy is a protective factor exploited by mutant HSCs to sustain disease and evade therapy may identify new therapeutic options to treat MPNs. Aims: The aim of the work is to provide a functional characterization of the biological relevance of dormant JAK2V617F-mutant HSCs in disease propagation and therapy resistance in a mouse model of PV. Methods: We have employed a novel transgenic mouse model that combines the knock-in of JAK2V617F mutation with the incorporation of the fusion protein histone H2B-GFP into the chromatin of HSCs. By using this model, which recapitulates a PV-like phenotype and allows the tracking of past replication history at a single HSC level, we have performed label-retention assays to formally interrogate the existence of long-term dormant mutant HSCs and their response to therapy. Results: Our results show that a rare subpopulation of mutant HSCs persists in a dormant state for up to 20 weeks post chase, despite the constitutively active JAK-STAT signaling. Moreover, our preliminary transplantation data supports the concept that dormant mutant HSCs may be responsible for disease propagation. Upon treatment with Fedratinib alone, dormant mutant HSCs persist, while treatment with a standard dose of Peg-IFN-α depletes these cells. Strikingly, while low dose Peg-IFN-α had no effect on dormant HSCs, the combination of Fedratinib and low-dose Peg-IFN-α synergized to deplete the dormant HSCs while also promoting a hematologic response. This suggests that low dose therapy with Peg-IFN-α can effectively deplete disease propagating HSCs when used in concert with Fedratinib, potentially circumventing the Peg-IFN-α-associated side effects. Summary/Conclusion: We have made the first characterization of dormant PV HSCs, that are able to maintain a quiescent state in vivo for more than four months, despite the presence of constitutive Jak2 signaling. Our preliminary data also indicates that these cells are relevant for disease propagation and that they are resistant to pharmacologic inhibition of Jak2 signaling. We hypothesize that dormancy might represent a state where mutant HSCs are no longer reliant on constitutive Jak2 signaling for survival. Importantly, combination treatment with Fedratinib and low-dose Peg-IFN-α produced a molecular response equivalent to that achieved using the standard dose of Peg-IFN-α, suggesting that low dose Peg-IFN-α can render these dormant HSCs again reliant on JAK2V617F signaling. We are currently using this model to establish how the status of cellular dormancy is maintained in a sub-set of HSCs in the face of constitutive JAK2 signaling, since this may identify additional therapeutic strategies to target this therapy-resistant cell population. Keywords: Interferon alpha, Hematopoietic stem cell, Myeloproliferative disorder